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BMRB: 36385

7D2O

Solution structure of Gaussia Liciferase by NMR

Authors

Kobayashi, N., Wu, N., Kuroda, Y., Yamazaki, T.

Assembly

Luciferase

Entity

1. Luciferase (polymer, Thiol state: all disulfide bound), 174 monomers, 18824.60 Da Detail

TGKPTENNED FNIVAVASNF ATTDLDADRG KLPGKKLPLE VLKEMEANAR KAGCTRGCLI CLSHIKCTPK MKKFIPGRCH TYEGDKESAQ GGIGEAIVDI PAIPRFKDLE PMEQFIAQVD LCVDCTTGCL KGLANVQCSD LLKKWLPQRC ATFASKIQGQ VDKIKGAGGD IEGR

Formula weight

18824.6 Da

Entity Connection

disulfide 5 Detail

ID	Type	Value order	Atom ID 1	Atom ID 2
1	disulfide	sing	1:CYS54:SG	1:CYS129:SG
2	disulfide	sing	1:CYS58:SG	1:CYS125:SG
3	disulfide	sing	1:CYS61:SG	1:CYS122:SG
4	disulfide	sing	1:CYS67:SG	1:CYS79:SG
5	disulfide	sing	1:CYS138:SG	1:CYS150:SG

Source organism

Gaussia princeps

Exptl. method

Refine. method

simulated annealing

Data set

assigned_chemical_shifts

Chem. Shift Complete

Sequence coverage: 97.1 %, Completeness: 89.8 %, Completeness (bb): 95.7 % Detail

Polymer type: polypeptide(L)

	Total	¹H	¹³C	¹⁵N
All	89.8 % (1781 of 1983)	87.8 % (909 of 1035)	91.2 % (695 of 762)	95.2 % (177 of 186)
Backbone	95.7 % (982 of 1026)	94.9 % (338 of 356)	96.2 % (486 of 505)	95.8 % (158 of 165)
Sidechain	85.5 % (952 of 1114)	84.1 % (571 of 679)	87.4 % (362 of 414)	90.5 % (19 of 21)
Aromatic	97.7 % (86 of 88)	97.7 % (43 of 44)	97.7 % (42 of 43)	100.0 % (1 of 1)
Methyl	96.8 % (180 of 186)	96.8 % (90 of 93)	96.8 % (90 of 93)

1. entity 1

TGKPTENNED FNIVAVASNF ATTDLDADRG KLPGKKLPLE VLKEMEANAR KAGCTRGCLI CLSHIKCTPK MKKFIPGRCH TYEGDKESAQ GGIGEAIVDI PAIPRFKDLE PMEQFIAQVD LCVDCTTGCL KGLANVQCSD LLKKWLPQRC ATFASKIQGQ VDKIKGAGGD IEGR

Show sample condition

Sample

Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

Protein Blocks Logo

Calculated from 19 models in PDB: 7D2O, Strand ID: A Detail

Release date

2020-11-29

Citation

Solution structure of Gaussia Luciferase with five disulfide bonds and identification of a putative coelenterazine binding cavity by heteronuclear NMR
Wu, N., Kobayashi, N., Tsuda, K., Unzai, S., Saotome, T., Kuroda, Y., Yamazaki, T.
Sci. Rep. (2020), 10, 20069-20069, PubMed 33208800 , DOI 10.1038/s41598-020-76486-4 , Abstract

Related entities 1. Luciferase, : 1 : 1 entities Detail

Experiments performed 8 experiments Detail

1 2D 1H-15N HSQC

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

2 2D 1H-13C HSQC aliphatic

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

3 2D 1H-13C HSQC aromatic

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

4 3D HNCO

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

5 3D HNCACB

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

6 3D HN(CO)CA

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

7 3D 1H-15N NOESY

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

8 3D 1H-13C NOESY

Instrument

Bruker AVANCE III - 700 MHz

Sample

State isotropic, Solvent system 90% H2O/10% D2O, Pressure 1 atm, Temperature 293 K, pH 6.0, Details 200 uM [U-13C; U-15N] Gluc, 50 mM MES, 2 mM sodium azide, 90% H2O/10% D2O

#	Name	Isotope labeling	Type	Concentration
1	Gluc	[U-13C; U-15N]	protein	200 uM
2	MES	natural abundance	buffer	50 mM
3	sodium azide	natural abundance		2 mM
4	H2O	natural abundance	solvent	95 %
5	D2O	[U-2H]	solvent	5 %

Chemical shift validation 3 contents Detail

Properties

Input source #1: Assigned chemical shifts - Assigned chemical shifts	bmr36385_3.str
Diamagnetism of the molecular assembly	True (excluding Oxygen atoms)
Whether the assembly has a disulfide bond	True (see coordinates for details)
Whether the assembly has a other bond	None
Whether the assembly contains a cyclic polymer	None
Overall data processing status	Warning

Disulfide bonds

Ptnr_site_1	Ptnr_site_2	Redox_state_prediction_1	Redox_state_prediction_2	Distance (Å)
1:54:CYS:SG	1:129:CYS:SG	oxidized, CA 53.9, CB 41.505 ppm	oxidized, CA 58.701, CB 39.066 ppm	n/a
1:58:CYS:SG	1:125:CYS:SG	oxidized, CA 58.635, CB 38.628 ppm	oxidized, CA 53.574, CB 40.772 ppm	n/a
1:61:CYS:SG	1:122:CYS:SG	oxidized, CA 60.959, CB 39.266 ppm	oxidized, CA 51.652, CB 35.27 ppm	n/a
1:67:CYS:SG	1:79:CYS:SG	oxidized, CA 56.257, CB 45.762 ppm	oxidized, CA 54.337, CB 43.228 ppm	n/a
1:138:CYS:SG	1:150:CYS:SG	oxidized, CA 56.49, CB 42.283 ppm	oxidized, CA 54.749, CB 40.667 ppm	n/a

Other bonds (neither disulfide, covalent nor hydrogen bonds, e.g. Zinc–sulphur bond)

None

Non-standard residues

None

Content subtype: bmr36385_3.str

#	Content subtype	Saveframe	Status	# of rows	Experiment type	Sequence coverage (%)
1	Assigned chemical shifts	assigned_chemical_shifts_1	Warning	1960		96.6 (chain: 1, length: 174)
1	Chemical shift references	chem_shift_reference_1	OK	3		No information

Assigned chemical shifts

Saveframe: assigned_chemical_shifts_1
- Status: Warning
- Sequence coverage of experimental data
  - Entity_assembly_ID: 1, Chain length: 174, Sequence coverage: 96.6%
- Total number of assignments
  - ¹H chemical shifts: 1094
  - ¹³C chemical shifts: 689
  - ¹⁵N chemical shifts: 177
- Completeness of assignments
  - Entity_assemble_ID: 1
- Redox state of CYS
- Peptide bond of PRO
- Tautomeric state of HIS
- Rotameric state of ILE , LEU, and VAL
- Histogram of normalized assigned chemical shifts
- Random coil index and derived parameters (S² and NMR RMSD)
  - Chain_ID: 1

Keywords LUMINESCENT PROTEIN, Luciferase